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Treatment of cancer often requires exposure to radiation, which has several limitations involving non-specific toxicity toward normal cells, reducing the efficacy of treatment. Efforts are going on to find chemical compounds which would effectively offer protection to the normal tissues after radiation exposure during radiotherapy of cancer. In this regard, plant-derived compounds might serve as “leads” to design ideal radioprotectors/radiosensitizers. This article reviews some of the recent findings on prospective medicinal plants, phytochemicals, and their analogs, based on both in vitro and in vivo tumor models especially focused with relevance to cancer radiotherapy. Also, pertinent discussion has been presented on the molecular mechanism of apoptotic death in relation to the oxidative stress in cancer cells induced by some of these plant samples and their active constituents.

Introduction

Cancer is now the third leading cause of death worldwide, with an estimated 12 million new cases and 7.6 million cancer deaths reported in American Cancer Society (2007). It is projected that by 2030 there would be about 26 million new cases, while a majority of deaths due to cancer will occur in developing countries (Bray and Moller, 2006; World Cancer Report, 2008). In the meantime, the global distribution of cancer along with the predominating types of the disease continues to change. Thus, cancers of the lung, breast, colon/rectum, and prostate are no longer confined to the Western industrialized countries but are among the most common cancers occurring all over the world. As for the therapy and management of cancer, newer strategies comprising multi-faceted and integrative approach involving surgery, followed by chemotherapy along with radiation is currently gaining consensus (Oehler et al., 2007). At the same time, advancement in the understanding of the disease processes at molecular level has offered novel targets for prevention, detection, control, and elimination of cancer.

Application of ionizing radiation, over and above surgery, and chemotherapy, has been the treatment of choice in case of solid malignancies (Kinsella, 2011). However, a substantial fraction of such tumors would fail to respond well to the radiation treatment, and require a very high dose to get killed, posing a severe limitation to the radiotherapy. Additionally, undesirable complications would occur owing to radiation injury to the surrounding normal tissues and to the skin, brain, heart, lung, kidney, liver, or gastrointestinal system of the cancer patient. Also, symptoms like tissue fibrosis, hair loss, xerostomia, xerophthalmia, etc., considerably restrict the application of a high dose of radiation aimed at the tumor-bearing organs (Dest, 2006).

Moreover, co-administration of radiation (delivered in the range of 40–80 Gy) along with the chemotherapeutic regimen might aggravate these complications (Curry and Curran, 2003). Patients undergoing treatment with taxol or vincristine often suffer from peripheral neuropathy as a side effect, while anthracycline drugs like doxorubicin (adriamycin), epirubicin, and mitoxantrone, etc., might lead to cardiac dysfunction (Choy, 2001). Abnormal kidney function and hearing loss were some of the common adverse effects occurring upon radiotherapy given to patients under treatment with platinum compounds (Amorino et al., 2000). Combination with some of the alkylating agents, like cyclophosphamide, ifosfamide, and leukeran (chlorambucil), might cause infertility (Verma et al., 2007). Again, a long term problem might emerge due to the post-radiotherapy incidence of a second tumor appearing either at the site of irradiation or away from it (Ng et al., 2002). Hence, it is a major challenge to radiation oncologists and researchers to develop alternative approaches to minimize the dosage through selective sensitization of tumor cells to respond to the radiation treatment, and thereby evade the detrimental consequences of radiotherapy (Rosenberg and Knox, 2006).

The exposure to radiation would primarily generate intracellular reactive oxygen species (ROS, viz., superoxide and hydroxyl radicals), which in turn would lead to DNA strands breaks and conformational alterations of biomolecules (Halliwell and Gutteridge, 1989). This will inevitably cause damage to surrounding normal cells. Hence, certain compounds/formulations could be envisaged to effectively scavenge the free radicals and thereby protect the surrounding normal cells from radiation induced injury. Historically, the seminal findings on the radioprotective ability of naturally occurring amino-metabolites like cysteine and cysteamine triggered the search for other thiolamines which would protect us from the acute effects of radiation (Patt et al., 1949). Thus, amifostine ([S-2-[3-aminopropylamino] ethylphosphorothioic acid) was developed as a potential radioprotector molecule (Weiss and Landauer, 2000; Grdina et al., 2002; Bensadoun et al., 2006). Nevertheless, its wider applicability has been restricted due to the major limitations associated with nausea, diarrhea, hypotension, hypocalcaemia, sleeplessness, dizziness, nephro- and neuro-toxic effects. Other non-protein thiols possessing antioxidative properties, viz. captopril ([S]-1-[3-mercapto-2-methyl-1-oxo-propyl]-L-proline), mesna (sodium-2-mercapto-ethanesulfonate), and N-acetyl-L-cysteine (NAC), were also relevant in this regard (Murley et al., 2004). However, these exogenous antioxidants proved to be ineffective if administered at the post-irradiation stage. These scavengers, acting only for a limited period of time (15 min to 1 h under in vivo conditions), must be administered shortly before the radiation exposure as the generation of highly reactive free radicals following radiation is a very rapid process, spanning less than 10−3 s (Grdina et al., 2002). Hence, there is an urgent need to look for more suitable molecules to be used in combination with chemo- and radio-therapy of cancer in order to minimize the adverse effects, and to enhance the overall curative outcome in the patients.

Screening and testing of compounds from natural as well as synthetic sources have been carried out over the last few decades in order to find effective radioprotectors capable of inhibiting radiation damage not only during radiotherapy of cancer patients, but also to healthy individuals undergoing occupational and accidental exposures to radiation (Stone et al., 2003). In this context, several authors have reviewed the prospective application of traditional medicinal plants which are known to contain anti-inflammatory, antioxidant, and immunomodulatory compounds (Arora et al., 2005; Venkatachalam and Chattopadhyay, 2005; Jagetia, 2007). Again, plant-derived polyphenolic compounds with radiosensitizing property have been extensively reviewed elsewhere (Garg et al., 2005). Incidentally, some of these antioxidants and plant products were also shown to be effective in prevention of cancer incidence (Suresh and Vasudevan, 1994; Zhao et al., 1997; Lee et al., 2002; Girdhani et al., 2005). The present article would be following up these developments during the last 5 years, and aim to highlight the current endeavor (since 2006) to identify phytochemicals and secondary metabolites of medicinal plants with relevance to cancer radiotherapy, and at the same time, attempt to elucidate the mechanistic premise in the light of available reports.

Potential Plant Products for Application in Cancer Radiotherapy

The global search for naturally occurring phytochemicals as potential radiotherapeutic agents has unearthed a host of plant products broadly categorized as (i) “radioprotectors”- to ameliorate the undesired damages caused to the normal cells, hence, minimize the side effects of radiation therapy; and (ii) “radiosensitizers”- to enhance the radiation-induced cell death inflicted to the tumor, and thereby minimize the dose of radiation treatment. In the present article, some of the major findings on traditional medicinal plants and active phytochemicals with promising radioprotective or radiosensitizing efficacy have been briefly summarized in a tabular form (Tables 1 and 2; post-2006).

Table 2. Phytochemicals with prospective radioprotective/radiosensitizing efficacy: reports from last 5 years study.

In Table 1, we have enlisted the reports on the crude extracts/semi-purified fractions of plant samples which demonstrated substantial prospect to enhance the clinical success of radiotherapy through a combination treatment. It is to be noted that 14 out of the twenty six plants in this list, viz. Aloe arborescens, Angelica sinensis, Azadirachta indica, Biophytum sensitivum, Boerhaavia diffusa, Citrus sinensis, Genista sessilifolia, Grewia asiatica, Isatis indigotica, Moringa oleifera, Olea europaea, Rosmarinus officinalis, Rubus spp., and Xylopia aethiopica have not been investigated prior to 2006. Most of the reports were obtained from the in vivo study on mouse models, where the radio-protecting activity was found to be associated with significant scavenging of free radicals, and depletion in lipid peroxidation with elevation in the glutathione, catalase, and lactate dehydrogenase enzyme levels. Also, several of these studies were conducted on propagatory cell lines and primary cultures to unravel the underlying mode of action at the molecular level (Table 1; Kimura and Sumiyoshi, 2009; Lee et al., 2010; Park et al., 2011).

In Table 2, we have presented the pure plant constituents and/or their analogs which could be considered as emerging candidates to be developed for the aforesaid application in future. Here, it is to be noted that the radiotherapeutic prospect of several plant-derived compounds, viz. allicin, betulinic acid, crocetin, diospyrin, honokiol, maytansine, oleuropein, α-santalol, tangeritin, withaferin A, and zingerone have been reported for the first time during the last 5 years.

Here, in Table 2, it has to be mentioned that we have not included the established plant-derived anticancer drugs, viz. etoposide, pactitaxel, and Vinca alkaloids, which have not only been recognized as potential radiosensitizers, but already under clinical application in association with cancer radiotherapy (Burris and Hurtig, 2010). Nevertheless, these drugs and their analogs are also under continuous appraisal for further development in this regard (Hiro et al., 2010; Orditura et al., 2010; Lillo et al., 2011; Schwarzenberger et al., 2011).

Again, prospective radiotherapeutic application of herbal formulations composed of traditional medicinal plants, and marketed as Triphala, Abana, Mentat, Septilin, Chyavanaprasha, Oligonol, HemoHIM, Fuzheng zengxiao formula, etc., have been reported by Sandhya et al. (2006), Jagetia (2007), Kundu et al. (2008), Park et al. (2010), Huang et al. (2011). In fact, the potent radioprotective property of Triphala, a mixture of three plants, might actually be attributed to the presence of Emblica officinalis (vide Table 1), which is also a major constituent of some of the other Oriental rejuvenators (Chyavanaprasha, Septilin, etc.) found to offer protection against radiation damage. Likewise, some herbal products, like HemoHIM from Far-East countries, contain rhizhomes of Angelica spp., a prospective source of radioprotectants (vide Table 1), while Oligonol is composed of modified plant phenolics. Therefore, these commercial formulations comprising poly-herbal mixtures have been kept outside the purview of the present article in order to focus on the search for new plants and phytochemicals with prospective radiotherapeutic property, and not included in our Tables.

Molecular Mechanism of Natural Radioprotectors/Radiosensitizers

Over the years, multi-modal therapy involving more than one anticancer agent applied in combination has been found to be favorable in the management of cancer. The precise efficacy and degree of tumor control exhibited by combination regimen, however, remains variable. Although the reasons for variability remain unclear, discovery of additional novel drugs that synergize with an existing radiation therapy would allow multiple combinations to choose from, thereby increasing the likelihood of clinical success. Recently, Edwards et al. (2011) has developed an interesting Drosophila larvae model which could be used to screen and identify molecules that would act in conjunction with radiation therapy (Edwards et al., 2011). On the whole, a number of phytochemicals with anti-/pro-oxidant, or immunomodulatory activity hold greater promise in pre-clinical/clinical trials. Emerging data also demonstrated that many phytochemicals, especially camptothecin, epigallocatechin gallate (EGCG), paclitaxel, etoposide, curcumin, etc., have potent growth inhibitory and apoptosis inducing effects on human as well as animal cancer cells by targeting multiple cellular signaling pathways in vitro. Therefore, these compounds could be useful in combination with conventional chemotherapeutic agents/radiation for the treatment of cancer, and expected to have lower toxicity but higher effectiveness. Also, recent in vivo pre-clinical studies and clinical trials have provided increasing evidence in support of multi-targeted therapies in combination with natural products. A comprehensive view on the molecular mechanisms to rationalize the prospective role of such phytochemicals acting on relevant signaling pathways has been given in Table 2.

Scavenging of Reactive Oxygen Species

Natural products in conjunction with irradiation are likely to exert the protective action through several mechanisms. Scavenging of free radicals generated during radiolysis would be a credible mode of action. Hence, naturally occurring polyphenolic compounds and antioxidant vitamins, primarily retinoids, would be the plausible candidates to offer radio-protection. It is a fact that the chances of developing cancer could be minimized through optimum nutritional supplementation by consuming a variety of fruits and vegetables, some of which have displayed chemopreventive activity by inhibiting tumorigenesis induced by chemical carcinogens and other genotoxic agents (Loo, 2003).

However, clinical reports on application of plant extracts with antioxidant property as adjuvants in cancer radiotherapy are still sparse in literature. Apparently, there is an apprehension that the antioxidants would protect not only the normal cells, but also the tumors, from the attack of free radicals generated during the course of treatment with ionizing radiation and anticancer agents. The lack of strong experimental evidences to address this concern resulted in poor enthusiasm from radiation oncologists to recommend their patients to consume such antioxidant products during the course of therapy. However, the pros and cons of this aspect have been critically reviewed, based on in vitro and in vivo experimentations (Prasad, 2005; Prasad and Cole, 2006).

In this context, hydrogen peroxide (H2O2) has been known to play a crucial role in the proliferation of cancer cells. In fact, many human cancers, like melanoma, neuroblastoma, colon carcinoma, and ovarian carcinoma, were found to constitutively generate a high amount of H2O2 (Szatrowski and Nathan, 1991). This indicated that the tumor cells would require a certain level of oxidative stress for maintaining a balance to undergo either proliferation or apoptotic death, and a minor fluctuation in the concentration of ROS might be critical to the intracellular signaling mechanism (Droge, 2002). The production of a larger amount of H2O2 was demonstrated by transforming NIH3T3 cells with Ras oncogene to cancer cells (Benhar et al., 2001), and a similar observation was noted in our laboratory when thymus cells in mice were transformed to thymic lymphoma after whole body radiation exposure (Pandey et al., unpublished data). The elevated production of ROS in cancer cells might be routed through mitochondrial electron transport chain, peroxisomes or NAD(P)H oxidase pathways, but the involvement of some direct mechanism of H2O2 generation may also be suggested. The constitutive production of ROS caused sub-lethal DNA damage in tumors, which was evidenced by a higher level of 8-hydroxy-2′-deoxyguanosine, while the increase of 4-hydroxy-2-non-enal, the lipid peroxidation product, indicated damage in cell membrane of carcinoma tissues (Toyokuni et al., 1995; Kondo et al., 1999). The cancer cells, in spite of having some amount of sub-lethal DNA damage, are generally adapted to survive in such stress conditions, and do not undergo cell cycle arrest or apoptosis (Elledge and Lee, 1995). In fact, at the basal level, the ROS would provide a stimulatory environment conducive to the proliferation of cancer cells, and become somewhat vital to their survival, rather than being merely useless cytotoxic products. Therefore, it may be hypothesized that the phenolic compounds with antioxidant properties could induce cell cycle arrest and apoptosis through scavenging of H2O2, presumably by depriving the cancer cells of an essential factor vital to their sustenance. It could also be speculated that a relatively lower level of constitutive ROS in normal cells would make them less vulnerable to the phenolic antioxidants (Simone et al., 2007).

Generation of Reactive Oxygen Species

Another category of phytochemicals showing antitumor activity are those which would enhance the generation of ROS, instead of scavenging the cellular free radicals. Such ROS-generators would apparently sensitize cancer cells endowed with persistent oxidative stress to undergo apoptotic death. We have already discussed about the critical maintenance of constitutively produced ROS, which is probably just below the threshold level required to induce apoptosis in tumors, the basal level being much lower in case of normal cells (Droge, 2002). Therefore, ROS-generating quinones could presumably create the requisite imbalance to lead the tumors cells, rather than the normal ones, to apoptotic death. Thus, the pro-oxidant quinones like β-lapachone (Suzuki et al., 2006; Dong et al., 2010a,b), plumbagin (Nair et al., 2008), and diospyrin derivatives (Hazra et al., 2007; Kumar et al., 2007, 2008) have been found to induce apoptosis in tumor cells through DNA damage, lipid peroxidation, mitochondrial membrane depolarization, and related signaling events. Incidentally, a few other anticancer plant products, such as paclitaxel, vinca alkaloids, and maytansinol, have been found to enhance the effect of radiation in human cancer cells through the involvement of microtubule interference to inhibit the proliferation of tumor (Edwards et al., 2011).

In our laboratory, we are investigating the radiomodulating potential of a diethtylether derivative (D7) of diospyrin, an antitumor quinonoid plant-product, in human breast carcinoma cells (MCF-7). It was observed that D7, in combination with radiation, could increase the apoptosis in tumor cells through down-regulation of the anti-apoptotic Bcl-2 and COX-2 gene expression, and up-regulation of pro-apoptotic genes, like p53 and p21. The higher expression of PUMA (p53 upregulated modulator of apoptosis), a pro-apoptotic protein, was also observed in the combination treatment. Further, the up-regulation of p21 expression in irradiated MCF-7 cells was found to be concomitant with the cell cycle arrest in the G1 phase (Kumar et al., 2007). Further studies in mouse and human fibrosarcoma cells (viz., Wehi164 and HT1080, respectively) showed marked enhancement of cytotoxicity with decreased clonogenic survival following treatment with D7 in combination with radiation. Moreover, increased radiosensitivity of tumor cells by D7 was found to occur through inhibition of radiation-induced NF-κB activation with substantial generation of intracellular ROS, ultimately leading to programmed cell death. Further, a combination regimen of D7 with 5 Gy radiation administered in two fractionated doses (2.5 Gy each) could cause a significant inhibition of tumor growth and increased life span of experimental mice bringing the liver enzyme activity to the normal level (Kumar et al., 2008).

Conclusion

Cancer patients need to go through extensive treatment involving chemotherapy, surgical intervention, recurring exposure to gamma-irradiation, or a combination therapy. Some traditionally popular medicinal plants have recently gained attention for their ability to modulate a number of signaling pathways that could initiate and facilitate the proliferation of cancer. In many cases, the potency of these compounds/formulations to sensitize the cancer cells to radiotherapy could be corroborated with the inhibition/activation of the relevant molecular markers. However, the literature citations on supporting clinical trials showing similar observations are quite limited. Nevertheless, a number of reports are available on antioxidants being able to protect against radiation-induced oncogenic transformations in experimental systems. Based on these information it has been presumed that supplementation of vitamins in a good measure, and intake of health promoting plant products in the diet might reduce the harmful side effects of standard therapeutic modalities and enhance their selective toxicity toward malignant cells, leading to an overall improvement in the efficacy of anticancer treatment.

Furthermore, the underlying mechanism of survival and proliferation in some types of cancer would reveal the inherent dependence of these cells on their constitutive oxidative stress. This mechanistic interpretation, in the light of the well-studied role of plant-polyphenolics in scavenging cellular free radical species, might resolve the prevailing dilemma on whether antioxidants would provide the desirable relief, to some extent, to the normal cells in preference to the tumor-bearing ones. Again, this hypothesis would be relevant to the radiosensitizing effect exhibited by a few ROS-generating antitumor agents as well. Thus, it is hoped that future research would add up positively, and would bring more of the aforesaid phytochemicals from “bench to bedside” of the suffering humanity seeking relief from the awful maladies of cancer.

Conflict of Interest Statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Acknowledgments

Banasri Hazra is grateful to the Board of Radiation and Nuclear Sciences, Department of Atomic Energy, Mumbai (Grant No. 2008/37/30/BRNS), and the University Grants Commission, New Delhi, for financial support. Subhalakshmi Ghosh is a Research Associate under this program.